Analysis of the effects of ß-mannanase on immune function and intestinal flora in broilers fed the low energy diet based on 16S rRNA sequencing and metagenomic sequencing.

As an enzyme, ß-mannanase (BM) can be widely used as feed additive to improve the growth performance of animals. This experiment aimed to determine the effect of the addition of BM to low-energy diet on the immune function and intestinal microflora of broiler chickens. In this study, 384 one-day-old Arbor Acres broilers were randomly divided into 3 groups (8 replicates per group): positive control (PC, received a corn-soybean meal basal diet), negative control (NC, received a low-energy diet with Metabolizable Energy (ME) reduced by 50 kcal/kg) and NC + BM group (NC birds + 100 mg/kg BM). All birds were raised for 42 d. The results showed that BM mitigated the damage of immune function in peripheral blood of broilers caused by the decrease of dietary energy level by increasing the Concanavalin A (Con A) index of stimulation (SI) and macrophages phagocytic activity in the peripheral blood of broilers at 42 d (P < 0.05). The analysis of cecum flora showed that the low-energy diet significantly reduced the observed_species index (P < 0.01), Chao1 index and ACE index (P < 0.05), which reduced the abundance and evenness of species in the cecum of broilers at 21 d. It also significantly reduced the relative abundance of Candidatus_Arthromitus and significantly increased the relative abundance of Pseudomonas in the cecum of broilers at 21 d, while also significantly increasing the relative abundance of Monoglobus at 42 d. BM significantly increased the relative abundance of Lachnospiraceae_UCG-001 and Lachnospiraceae_bacterium_615 in the cecum of broilers at 21 d. In addition, BM inhibited microbial Fatty acid degradation by decreasing the activity of glutaryl-CoA dehydrogenase. Collectively, BM could improve intestinal health by enhancing the immune function of broilers, promoting the proliferation of beneficial bacteria and reducing the number of harmful bacteria, regulating intestinal flora, thereby alleviating the adverse effects of lower dietary energy levels.

Probiotic cocktails accelerate baicalin metabolism in the ileum to modulate intestinal health in broiler chickens.

BACKGROUND: Baicalin and probiotic cocktails are promising feed additives with broad application prospects. While probiotic cocktails are known to enhance intestinal health, the potential synergistic impact of combining baicalin with probiotic cocktails on the gut health of broiler chickens remains largely unexplored. Therefore, this study aims to investigate the influence of the combined administration of baicalin and probiotic cocktails on the composition of ileal and cecal microbiota in broiler chickens to elucidate the underlying mechanisms responsible for the health-promoting effects. RESULTS: A total of 320 1-day-old male Arbor Acres broilers were divided into 4 groups, each with 8 replicates of 10 chicks per replicate. Over a period of 42 d, the birds were fed a basal diet or the same diet supplemented with 37.5 g/t baicalin (BC), 1,000 g/t probiotic cocktails (PC), or a combination of both BC (37.5 g/t) and PC (1,000 g/t). The results demonstrated that BC + PC exhibited positive synergistic effects, enhancing intestinal morphology, immune function, and barrier function. This was evidenced by increased VH/CD ratio, sIgA levels, and upregulated expression of occludin and claudin-1 (P < 0.05). 16S rRNA analysis indicated that PC potentiated the effects of BC, particularly in the ileum, where BC + PC significantly increased the α-diversity of the ileal microbiota, altered its ß-diversity, and increased the relative abundance of Flavonifractor (P < 0.05), a flavonoid-metabolizing bacterium. Furthermore, Flavonifractor positively correlated with chicken ileum crypt depth (P < 0.05). While BC + PC had a limited effect on cecal microbiota structure, the PC group had a very similar microbial composition to BC + PC, suggesting that the effect of PC at the distal end of the gut overshadowed those of BC. CONCLUSIONS: We demonstrated the synergistic enhancement of gut health regulation in broiler chickens by combining baicalin and probiotic cocktails. Probiotic cocktails enhanced the effects of baicalin and accelerated its metabolism in the ileum, thereby influencing the ileal microbiota structure. This study elucidates the interaction mechanism between probiotic cocktails and plant extract additives within the host microbiota. These findings provide compelling evidence for the future development of feed additive combinations.

The accessible chromatin landscape of lipopolysaccharide-induced systemic inflammatory response identifying epigenome signatures and transcription regulatory networks in chickens.

Lipopolysaccharide (LPS) can induce systemic inflammatory response (SIR) in animals. Understanding the regulatory mechanism of SIR and therapies to ensure healthy growth is urgently needed. Chromatin remodeling plays a crucial role in the expression of genes involved in immune diseases. In the present study, the ATAC-seq analysis revealed 3491 differential open chromatin sites in the spleen of chicks with SIR induced by LPS challenge, and we presented the motifs on these sites and the associated transcription factors. The regulatory network was presented by combining the differential open chromatin data with the mRNAs and exploded cytokines. Interestingly, the LPS challenge could regulate the mRNA expression of 202 genes through chromatin reprogramming, including critical genes such as TLE1 and JUN, which regulate signaling pathways such as I-κB kinase/NF-κB, Toll-like receptor, and downstream cytokine genes. Furthermore, dietary daidzein could inhibit DNA topoisomerase II, which reprograms the spatial conformation of chromatin in the inflammatory response and attenuates SIR. In conclusion, we successfully identified key genes directly regulated by chromatin reprogramming in SIR and demonstrated the chromatin epigenome signatures and transcriptional regulatory network, which provides an important reference for further research on avian epigenetics. There is great potential for alleviating SIR using dietary daidzein.

Effects of maternal hawthorn-leaf flavonoid supplementation on the intestinal development of offspring chicks.

Metabolic disorders in maternal generation during the late egg-laying period have adverse effects on neonatal development. The study was conducted to clarify the effects of maternal feeding of hawthorn-leaf flavonoid (HF) on the microbial community and intestinal development of chicks. Breeder hens were fed a basic corn-soybean diet, while the treatment groups were supplemented with 30 or 60 mg/kg HF. The offspring chicks were divided into CON, LHF, and HHF groups according to the maternal treatments. Maternal HF supplementation at 60 mg/kg increased the average daily gain and decreased the feed conversion rate of chicks (P < 0.05), but did not affect the average daily feed intake. HF treatments increased the villus height to crypt depth ratio and up-regulated the protein expressions of PCNA, IGF-1R, PI3K and p-mTOR in the jejunum (P < 0.05) of 1-day-old and 14-day-old chicks. Additionally, maternal HF treatment up-regulated the mRNA expression of tight junction transmembrane proteins (occludin) and scaffolding proteins (ZO-1 and ZO-2) in the jejunum of 1-day-old chicks (P < 0.05). Moreover, the maternal effects of HF on ZO-1 expression could last for 14 d (P < 0.05). Interestingly, dietary HF supplementation altered the vertically transmitted microbial community from breeder hens to chicks, especially increased the relative abundance of probiotics (i.e., Clostridium_sensu_stricto_1) in the meconium of chicks (P < 0.05), which may help with early gut microbiota colonization and intestinal development. In summary, dietary HF supplementation for breeder hens altered the bacterial community of neonates and might promote intestinal development of chicks through the IGF-1R/AKT/mTOR signaling pathway.

Methionine Source and Level Modulate Gut pH, Amino Acid Transporters and Metabolism Related Genes in Broiler Chickens.

This study determined the effects of two methionine (Met) sources at three total sulfur amino acids (TSAA) to lysine ratios (TSAA/Lys) on gut pH, digestive enzyme activity, amino acid transporter expression, and Met metabolism of broilers. The birds were randomly assigned to a 2 × 3 factorial arrangement with Met sources (dl-Met and dl-2-hydroxy-4-(methylthio)-butanoic acid (OH-Met)) and TSAA/Lys (0.58, 0.73, and 0.88) from 1 to 21 days. The results demonstrated that dl-Met and OH-Met supported the same growth performance, but high TSAA/Lys ratio reduced the feed intake and body weight (P < 0.05). OH-Met reduced the crop chyme pH and enhanced the jejunal lipase activity (P < 0.05). ATB0,+ expression decreased with increased dl-Met levels in the duodenum; the low TSAA/Lys ratio induced a stronger mRNA expression of basolateral Met transporters. OH-Met resulted in an increase of cystathionine ß-synthase expression in the liver and a decrease in serum homocysteine levels at middle TSAA/Lys ratio compared with dl-Met treatment (P < 0.05). In conclusion, two Met sources support the same growth, but OH-Met acidified the crop chyme. The investigated transporter transcripts differed significantly along the small intestine. At the middle TSAA/Lys ratio, OH-Met showed a higher metabolic tendency of the trans-sulfuration pathway compared with dl-Met.

Maternal supplementation with mulberry-leaf flavonoids improves the development of skeletal muscle in the offspring of chickens.

The development of skeletal muscle is a crucial factor in determining the meat yield and economic benefits of broiler production. Recent research has shown that mulberry leaves and their extracts can be used to significantly improve the growth performance of livestock and poultry. The present study aims to elucidate the mechanisms involved in the regulation of skeletal muscle development in broiler offspring by dietary mulberry-leaf flavonoids (MLF) supplementation from the perspective of maternal effect theory. A total of 270 Qiling broiler breeder hens were randomly assigned to 3 treatments with different doses of MLF (0, 30, 60 mg/kg) for 8 weeks before collecting their fertilized eggs. The chicken offspring at 13 and 19 d of embryonic stage, and from 1 to 28 d old after hatching were included in this study. The results showed that maternal supplementation increased the breast muscle weight and body weight of the offspring at the embryo and chick stages (P < 0.05). This was followed by increased cross-sectional area of pectoral muscle fibres at 14 d (P < 0.05). Further determination revealed a tendency towards increased serum levels of insulin-like growth factor 1 (IGF-1) (P = 0.092) and muscle fibre count (P = 0.167) at 1 d post-hatching following maternal MLF treatment, while serum uric acid (UA) was decreased at 14 d after hatching (P < 0.05). Moreover, maternal MLF supplementation significantly up-regulated the mRNA expression of the myogenic regulatory factor Myf5 in skeletal muscle at the both embryonic and growth stages (P < 0.05). The relative abundance of the downstream protein of BMPR2, Smad1 and p-Smad1/5/9 in the TGFß signalling pathway was significantly increased by maternal MLF treatment. Meanwhile, the increased expression of the target protein p-mTOR in the breast muscle of the offspring chicks is in accordance with the improved growth rate of the breast and the body. In conclusion, maternal MLF supplementation can promote muscle protein metabolism and muscle fibre development of chick embryos through upregulation of Myf5 expression and BMP/p-Smad1/5/9 axis, thereby improving growth performance of slow growing broiler.

Mannanase improves the growth performance of broilers by alleviating inflammation of the intestinal epithelium and improving intestinal microbiota.

This experiment aimed to discuss and reveal the effect and mechanism of mannanase on intestinal inflammation in broilers triggered by a soybean meal diet. In this experiment, 384 Arbor Acres broilers at 1 d old were randomly divided into 3 treatment groups. The broilers were fed a corn-soybean meal basal diet, a low-energy diet (metabolizable energy reduced by 50 kcal/kg), and a low-energy diet supplemented with 100 mg/kg mannanase for 42 d. The low-energy diet increased feed conversion ratio from 0 to 42 d, reduced ileal villus height and villus height-to-crypt depth ratio and upregulated the expression of nuclear factor kappa B (NF-κB) in the ileum (P < 0.05). It also reduced cecal short-chain fatty acids (SCFA), such as acetic acid (P < 0.05). Compared with low-energy diets, the addition of mannanase increased body weight at 42 d, promoted the digestibility of nutrients, and maintained the morphology and integrity of the intestinal epithelium of broilers (P < 0.05). In addition, mannanase upregulated the expression of claudin-1 (CLDN1) and zonula occludens-1 (ZO-1) in the jejunum at 21 d, downregulated the expression of ileal NF-κB, and increased the content of isobutyric acid in the cecum of broilers (P < 0.05). The results for the ileal microbiota showed that a low-energy diet led to a decrease in the relative abundance of Lactobacillus reuteri in the ileum of broilers. The addition of mannanase increased the relative abundance of Lactobacillus-KC45b and Lactobacillus johnsonii in broilers. Furthermore, a low-energy diet reduced the relative abundance of Butyricicoccus in the intestine of broilers and inhibited oxidative phosphorylation and phosphoinositol metabolism. Mannanase increased the relative abundance of Odoribacter, promoted energy metabolism and N-glycan biosynthesis, and increased the activities of GH3 and GH18. It is concluded that mannanase could improve the growth performance of broilers by reducing the expression of NF-κB in the ileum, increasing the production of SCFA in the cecum, suppressing intestinal inflammation, balancing the intestinal microbiota, reducing damage to the intestinal barrier, and improving the efficiency of nutrient utilization to alleviate the adverse effects caused by the decrease in dietary energy level.